1 Abstract 2 Modern molecular medicine has increasingly focused on developing novel target-specific molecular probes to 3 improve the prognosis and diagnosis of diseases and provide the best treatment regimens. To this end, in the 4 past two decades, nucleic acid aptamers have been developed and a series of aptamer-based methods and 5 materials have been used for biomarker identification and the development of targeted drugs. Aptamers are 6 single-stranded oligonucleotides composed of DNA or RNA, usually having a length of 15-60 nucleotides. They 7 demonstrate remarkable binding affinity to a variety of targets including intact cells. Compared to antibodies, 8 aptamers are generated through an in vitro selection process, termed SELEX (Systematic Evolution of Ligands by 9 Exponential Enrichment). Thus, it can be easily resynthesized and chemically modified, with promising batch-to- 0 batch consistency, easy modification and satisfying biocompatibility. It is worth noting that antibodies are still 1 among the most intriguing tools for molecular recognition. However, monoclonal antibodies took more than 20 2 years to be humanized since initial production and much longer for real clinical applications. In comparison, over 3 40 clinical trials with aptamers have already been conducted, highlighting their enormous potential in clinical 4 practice. 5 The PI?s lab has been a leading group in aptamer research and development. In the past decade, the Tan lab 6 has developed aptamer-based theranostics and biomarker discovery strategies. Ever since around ten years ago 7 we initiated the cell-SELEX technology to deliver DNA aptamers specifically targeting intact cells, we have now 8 developed more than 300 aptamers against many types of cancerous and other diseased cells. Besides, 9 combined with the cutting-edge nanomaterials, aptamer-based theranostics have been developed. Despite the 0 enormous potential and recent technical advances made with aptamers, their true impact in biomedicine has not 1 been fully realized. The gaps in advancing their utility are mainly in slow selection process, unknown binding 2 mechanism and low efficacy in using aptamers for clinic studies. The goals here are to address these 3 fundamental gaps and challenges, to push aptamers from molecules to medicine and to open new areas of 4 research using aptamers. 5 Out central goal here is dedicated to fulfilling the promise of aptamers as key molecular tools. Two major 6 areas of research will be carried out: 1) understanding the nature of aptamers and aptamer/target structure and 7 developing new strategies for their use as effective molecular probes and 2) using aptamers to discover and 8 pursue new avenues of research and development. Specifically, we will fill the gaps by developing one round 9 SELEX to increase efficiency and obtain robust aptamers. We will also use cryo-EM microscopy to understand 0 the structural information of selected aptamers with their targets for better-predicted biological performances. 1 Both will not be proposed in a regular NIH proposal as they will be viewed either as too ambitious or too 2 preliminary to support. In addition, we will also optimize and create new aptamers in such a way to show their full 3 utilities as molecular probes in molecular medicine: including aptamer based accurate and smart diagnosis and 4 precision therapy strategies.